An Ion Channel That Perpetuates Pain in Diabetic Neuropathy

Diabetes is on the rise, and consequently, so is the persistent and often intractable pain experienced by many diabetic patients with nerve injury. However, researchers still have a poor understanding of the source of that pain, making it difficult to design better treatments. But now, Peter McNaughton, King’s College London, UK, and colleagues have identified an ion channel that causes allodynia in diabetic mice, pointing to a possible therapeutic target in people with painful diabetic neuropathy (PDN).

Using the drug ivabradine, the researchers peripherally blocked all four types of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in mouse models of type 1 and type 2 diabetes. The drug reversed the hypersensitivity to touch that emerged in each model, without affecting pain thresholds in control mice. This analgesic effect likely resulted from inactivation of HCN2, as diabetic mice that were genetically modified to lack these channels showed normal pain responses.

“The authors make a very good case that HCN2 is essential for pain behavior in two diabetes models,” says Claudia Sommer, University of Würzburg, Germany, who was not involved in the study. “If the results translate to humans, this would mean that HCN2 is a new target for analgesia in painful diabetic neuropathy, and that pain in diabetes could be treated in the peripheral nervous system. Thus, drugs with central nervous system side effects would not be needed.”

The results were published September 27 in Science Translational Medicine.

The path to HCN2

HCN channels were first discovered in cardiac muscle cells, and eventually in neurons distributed throughout the nervous system. Strikingly, they conduct excitatory currents at hyperpolarized rather than depolarized membrane potentials, allowing cells that have reached the end of one action potential to fire again.

Because of these electrophysiological properties, researchers have long thought that HCN channels might be drivers of pathological pain (Chaplan et al., 2003). “There had been some indications, from 2003 onward, that blockers of HCN channels—drugs which block all four types indiscriminately—were quite effective analgesics,” said McNaughton.

Meanwhile, McNaughton’s own lab had started to make sense of how the small family of channels finely tunes the firing of sensory neurons, including nociceptors (Momin et al., 2008). And so, “I wondered which one of the four channels might be involved” in pain, he said.

In 2011, he and his colleagues found an answer. Knowing that HCN2 channels are expressed by more than half of nociceptors, the team genetically engineered mice missing only those channels, and only in pain-sensing neurons. Remarkably, unlike their normal littermates, the knockout animals developed neither inflammatory pain nor neuropathic pain, the latter modeled with chronic constriction injury (CCI) to the sciatic nerve (Emery et al., 2011; see related PRF news story).

“We found that HCN2 was the criminal, so to speak,” said McNaughton.

Even so, for Christoforos Tsantoulas, lead author of the new paper, the clinical relevance of the discovery was unclear. “Neuropathy due to disease is the main reason why people get chronic pain,” he said. And yet, drugs that enter clinical trials for the treatment of pain are rarely developed using disease models. Instead, they are usually evaluated in models of traumatic injury, such as the CCI model, he said.

To address this concern, Tsantoulas and his colleagues began with mice injected with streptozotocin (STZ), a model of type 1 diabetes. Within eight weeks, approximately 80 percent of the STZ-treated animals showed heightened responses to harmless touch, relative to animals that received a vehicle solution. And much like PDN patients, STZ mice had fewer nerve fibers innervating the skin, likely reflecting nerve damage.

When the researchers injected the STZ mice with a single dose of ivabradine, which cannot enter the central nervous system, they saw a decrease in mechanical hypersensitivity in the animals. Better still, eight doses delivered over four days completely reversed it.

Mice with a mutated version of the leptin receptor, a model of type 2 diabetes, responded similarly. That’s encouraging, said Tsantoulas, considering that type 2 diabetes occurs far more frequently than type 1.

“Ninety percent of diabetic patients have type 2 diabetes, but the research most people in the field do overwhelmingly uses a type 1 model,” according to Tsantoulas.

As for whether HCN2 in particular contributed to the observed hypersensitivity, and thus to the relief provided by ivabradine, the researchers turned to the knockout mice lacking HCN2 in nociceptors they had used in their 2011 study. Despite an increase in blood glucose levels and a decrease in nerve fiber density in response to STZ, there was no indication of allodynia in these animals.

The results from both the ivabradine and knockout experiments speak to the origin of pain in diabetic neuropathy, said Sommer. “The authors make a strong point that it’s peripherally generated.”

Bringing HCN2 into action

What allowed HCN2 to play a role in pain? In contrast to HCN1 and 3, HCN2 and 4 will open to a greater extent at the resting membrane potential if they bind cyclic adenosine monophosphate (cAMP). The authors reasoned, then, that diabetes might lead to a rise in intracellular cAMP levels, in turn shifting the voltage dependence of HCN2 activation.

To test this idea, they compared the amount of cAMP in the dorsal root ganglia (DRG) of mice that received STZ and developed allodynia to those that received a vehicle solution. They found that cAMP levels were eight times greater in the former compared to the latter. The expression of HCN2 itself, however, remained unchanged, as immunohistochemistry demonstrated that STZ and control mice had the same proportion of HCN2-positive DRG cells.

What about spontaneous pain?

While the study could help explain how diabetes lowers the threshold for painful touch, the disease can cause other sensory symptoms as well. In fact, different patients can feel different forms of pain, and among these, spontaneous pain is the most common (see related RELIEF feature story). Yet, “neither spontaneous pain behavior nor spontaneous activity in [pain-related fibers] was examined in the mouse models,” said Sommer. As a result, “we do not know if the models capture these features.”

McNaughton nonetheless thinks the current data make a compelling argument for the importance of HCN2 in spontaneous pain. “There was an elevation in the expression of c-Fos [a gene upregulated by neuronal activity] in lamina I and II of the spinal cord in STZ mice, which is almost certainly due to enhanced, ongoing input from nociceptive fibers,” he said. This c-Fos expression quickly disappeared after the researchers injected the mice with ivabradine.

“It’s not absolutely direct evidence, but we think, based on that evidence, that blocking HCN2 channels would be effective in reducing spontaneous pain as well as evoked pain in patients,” said McNaughton.

Ivabradine is already prescribed for chest pain and congestive heart failure, so could it be repurposed for PDN? McNaughton is doubtful, since it would affect not only HCN2 but also HCN4, which is present in the heart and acts as a pacemaker. That means only low doses of the drug would be safe for use in patients. “I wouldn’t be surprised if these doses are not that effective” for PDN, said McNaughton.

“What you really need to do is block HCN2 selectively,” he said. “From a clinical point of view, it’s a much more desirable way to go.”

Matthew Soleiman is a science writer residing in Nashville, Tennessee. Follow him on Twitter @MatthewSoleiman.